Then, 293T cells were transiently co-transfected with pBabe-Cul4A-myc-his and pCMV6-ANXA10-GFP (OriGene, Rockville, MD, USA) vectors using Lipofectamine 2000 transfection reagent (Invitrogen). Twenty-four hours after transfection, the cells were treated with 10 μg/mL of MG132 (Sigma) for 24 h, and then harvested in a NP-40 lysis buffer (150 mM NaCl, 50 mM Tris [pH 8.0], 1% NP40), protease inhibitor, and phosphatase inhibitor cocktail (Roche, Lewes, UK, USA). Immunoprecipitation was performed using the Catch and Release v2.0 Reversible Immunoprecipitation System (Millipore) according to the manufacturer’s protocols. Anti-GFP (OriGene) and Anti-Cul4A (Abcam) antibodies were used for immunoprecipitation.
Pbabe cul4a myc his
Manufactured by OriGene
Sourced in United States
PBabe-Cul4A-myc-his is a plasmid construct containing the Cul4A gene with myc and his tags. It is designed for expression studies in mammalian cells.
Automatically generated - may contain errors
Lab products found in correlation
Pcmv6 anxa10 gfp, by OriGene (2 mentions)
Protease inhibitor, by Roche (1 mentions)
Phosphatase inhibitor cocktail, by Roche (1 mentions)
Mg132, by Merck Group (1 mentions)
Anti gfp, by OriGene (1 mentions)
Catch and release v2.0 reversible immunoprecipitation system, by Merck Group (1 mentions)
Lipofectamine 2000 transfection reagent, by Thermo Fisher Scientific (1 mentions)
Anti cul4a, by Abcam (1 mentions)
Prk5 ha ubiquitin wt, by Addgene (1 mentions)
Anti ha tag antibody, by Cell Signaling Technology (1 mentions)
2 protocols using pbabe cul4a myc his
1
Investigating ANXA10-Cul4A Interaction
2
Annexin A10 Ubiquitination Regulation
Check if the same lab product or an alternative is used in the 5 most similar protocols
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The 293T cells were co-transfected with a combination of pBabe-Cul4A-myc-his and pCMV6-ANXA10-GFP (OriGene) with or without pRK5-HA-Ubiquitin-WT (Addgene, Cambridge, MA, USA). All the cells were treated with 10 µg/mL of MG132 for 24 h prior to lysis. Anti-GFP antibody was used for immunoprecipitation. Anti-HA tag antibody (Cell Signaling, Danvers, MA, USA) was used for the western blot analysis.
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